Approximately 16% of Americans aged 14-49 are currently seropositive for Herpes simplex virus type 2 (HSV- 2) and worldwide, 23 million new infections occur each year. Transmission of HSV-2 from mother to neonate at birth results in serious morbidity or death. HSV-2 infections are the leading cause of genital ulcer disease which increases the risk of acquiring other sexually transmitted infections including HIV. HSV vaccines that elicit systemic immune responses have failed in clinical trials. Preclinical studies n animals and humans suggest that virus-specific T cells at the site of genital HSV-2 infection are critical for protection. Genital tract- resident memory T cells resulting from previous HSV-2 infection have been shown to become activated within a few hours of HSV-2 re-challenge and rapidly clear the virus. A rational vaccine approach to induce these tissue-resident memory cells would be direct immunization of the genital tract but the vaginal mucosa does not have immune inductive sites and the multi-layered genital epithelium presents a barrier for most non-infectious vaccines. Alternative immunization sites have been tested with the intent of eliciting immune cells that home to the genital epithelium but this approach has not been uniformly successful. Expression of skin-homing integrins, mucosal integrins and CXCR3 chemokine receptor has been reported to play a role in trafficking of CD8+ T cells to the vaginal epithelium. However, trafficking of immune cells expressing these molecules to the genital tract is normally achieved in response to an inflammatory stimulus in the genital tract and many of the infiltrating inflammatory cells that also migrate to the inflamed site are potential targets for HIV. To induce specific trafficking of virus-specific CD8+ T cells to the vaginal epithelium in the absence of overt inflammation, we propose the use of two novel vaccine platforms in a prime and pull approach. The overall goals of these studies are to utilize a chemokine pull of vaccine-elicited CD8+ T cells into the genital epithelium to establish a population of genital tract-resident memory T lymphocytes and to determine the role of endogenous- and exogenous adjuvanting of the vaccines to increase expression of CXCR3 by vaccine-induced T cells thereby facilitating response to the chemokine pull signal. Our central hypothesis is that heterologous prime/boost immunization with our novel vaccine platforms adjuvanted with endogenous and exogenous ligands for pathogen pattern receptor (PRR) agonists will induce high numbers of CXCR3-expressing, virus-specific CD8+ T cells that will traffic to the vaginal mucosa in response to chemokine ligands. Aim one will determine the influence of individual PRR pathways induced by priming with our novel single cycle flavivirus vaccine on the magnitude, function, and CXCR3 expression of induced CD8+ T cells. Aim two will determine the role of PRR agonist adjuvants used with a synthetic vaccine platform on the magnitude, effector function, CXCR3 expression, and protection against HSV-2 challenge. Together, these studies will provide important information on the role of adjuvants and novel immunization regimens to protect against sexually transmitted viruses.